Direct inhibition of P/Q-type voltage-gated Ca^sup 2+^ channels by Gem does not require a direct Gem/Ca^sub v^[Beta] interaction

The Rem, Rem2, Rad, and Gem/Kir (RGK) family of small GTP-binding proteins potently inhibits high voltage-activated (HVA) Ca... channels, providing a powerful means of modulating neural, endocrine, and muscle functions. The molecular mechanisms of this inhibition are controversial and remain largely unclear. RGK proteins associate directly with Ca... channel β subunits (Ca...β), and this interaction is widely thought to be essential for their inhibitory action. In this study, we investigate the molecular underpinnings of Gem inhibition of P/Q-type Ca... channels. We find that a purified Gem protein markedly and acutely suppresses P/Q channel activity in insideout membrane patches, that this action requires Ca...β but not the Gem/Ca...β interaction, and that Gem coimmunoprecipitates with the P/Q channel α... subunit (Ca...α...) in a Ca...β-independent manner. By constructing chimeras between P/Q channels and Gem-insensitive low voltage-activated T-type channels, we identify a region encompassing transmembrane segments S1, S2, and S3 in the second homologous repeat of Ca...α... critical for Gem inhibition. Exchanging this region between P/Q and T channel Ca...α... abolishes Gem inhibition of P/Q channels and confers Ca...β-dependent Gem inhibition to a chimeric T channel that also carries the P/Q I-II loop (a cytoplasmic region of Ca...α... that binds Ca...β). Our results challenge the prevailing view regarding the role of Ca...β in RGK inhibition of high voltage-activated Ca... channels and prompt a paradigm in which Gem directly binds and inhibits Ca...β-primed Ca...α... on the plasma membrane. (ProQuest: ... denotes formulae/symbols omitted.)